Biological assembly

ABSTRACT

A biological assembly for effecting the examination of a sample in a liquid medium under magnification. The assembly includes a transparent slide with a sample chamber and an immediately adjacent overflow chamber formed on one main surface by a thin film. The sample chamber is receptive of a sample in a liquid medium. transparent slide, having a pattern for aligning it with the sample chamber, is disposed over the sample chamber. The transparent slide includes a grid etched into the lower surface thereof to enable the counting cells under magnification.

BACKGROUND OF THE INVENTION

The present invention is directed to a biological assembly and inparticular to a slide and slide cover for use in biological studies.

It is often necessary to examine biological material in order to makequantitative determinations relating thereto. For example, a semensample may require microscopic study to determine the sperm count in theliquid medium of the sample being observed.

Conventionally, there have been two methods for estimating the number ofcells in a suspension. One method uses a hemocytometer wherein thenumber of cells in a given volume is counted by direct microscopicexamination. Another method is the use of an electronic counter whereinthe cells of a given volume of a suspension are drawn through an orificeand registered electronically.

The hemocytometer comprises a slide and a precision ground cover slipwhich covers two ruled areas which are ruled in the form of grids. Whenthe cover slip is pressed down on the slide, a chamber is formedtherebetween. A drop of cell suspension is placed at the two edges ofthe cover slip so that the suspension flows into the chambers bycapillary action. Other chambers of this type are known as the HorwellChamber, the Makler chamber and the Hausser Chamber. These devices mustbe ground to great precision and thus are expensive to make and areaccordingly non-disposable. They are, therefore, cleaned after each useso that they can be reused.

U.S. Pat. No. 4,911,782 disclosed a biological slide assembly whichenables a fixed volume of sample and liquid medium to be received in achamber by capillary action in order to be observed under a microscope.The microscope must include a reference grid in the ocular, which isdivided into 100 squares, so that the cell count in a representative oneof the squares can be obtained by a human observer and used toapproximate the total number of cells within the grid. While this deviceis disposable, it is complex in design and difficult to use in practice.

SUMMARY OF THE INVENTION

The main object of the present invention is to overcome thedisadvantages of the prior art and to provide a biological assembly,which is simple in design, easy to use and inexpensive to make so thatit is easily disposable after use. These and other features andadvantages of the present invention are achieved in accordance with thepresent invention by a biological assembly for effecting the examinationof a sample in a liquid medium under magnification, comprising atransparent substrate having a first main surface, and means forming atleast one sample camber and an immediately adjacent overflow chamber onthe first main surface and receptive of a sample in a liquid medium,said means comprising a thin film having for each sample chamber andover chamber, means defining a first opening therein corresponding tothe sample chamber and means defining a second opening surrounding thefirst opening and corresponding to the overflow chamber, wherein thefirst and second openings form a separating wall of thin filmtherebetween.

The substrate of the biological assembly preferably is a slide,preferably made of transparent glass and which has a thin film ofphotoresist, epoxy or hydrophobic ink on the upper surface thereof.

The thin film forms a sample chamber which is preferably in the form ofa circular or elliptical cylinder having the height of the thin filmitself. Immediately adjacent to the sample chamber is an overflowchamber. The overflow chamber in one preferred embodiment is toroidaland coaxial to the sample chamber and separated by a cylindrical wall ofthin film. In another preferred embodiment the sample chamber iscircular or elliptical and the outer edge of the overflow chamber isrectangular or square.

The thin film is preferably from 5 to 75 μm in height, more preferably10, 20 or 50 μm. The sample chamber preferably has a volume of 2 to 15μl, and thus when the diameter of the circular sample chamber is 1.6 cmand the height is 20 μm, the volume is 4 μl.

In accordance with the present invention, each slide can have one or aplurality of sample chambers and associated overflow chambers disposedtherearound. Most preferably, the slide will have from one to threesample chambers and overflow chambers.

The biological assembly according to the present invention furthercomprises a slide cover having a substrate which is preferably made oftransparent glass and which is superposed on the slide to cover eachsample chamber and overflow chamber.

In accordance with the present invention, the slide cover preferably hasa pattern of thin film on the surface opposite that which covers thesample chamber, the pattern demarcating an area so as to enable thatarea to be aligned within the sample chamber. In a particularlypreferred embodiment of the present invention, the pattern on the slidecover is identical in configuration to that of the wall separating theoverflow chamber from the sample chamber.

The slide cover also preferably includes a grid formed in the surface ofthe slide cover which is closest to superposed on the sample chamber soas to be as close as possible to the sample in the liquid medium that isbeing observed. The grid is preferably 10×10 boxes of 0.1 mm on a side.The grid is disposed within the area demarcated by the pattern. In aparticularly advantageous embodiment of the invention, the grid isalmost invisible to the naked eye, so the pattern surrounding it is anindication to the user of the location of the grid.

In another embodiment of the present invention, the slide cover can beused with conventional slides, that is, slides other than that of thepresent invention. In a particularly advantageous embodiment of thisinvention, the slide cover has a thin film pattern on one main surfacedemarcating an area and spacing means composed of thin film on thesurface of the slide cover which faces the slide to space the grid by afixed distance form the upper surface of the slide to enable the grid tobe in contact with a sample which is disposed on the slide surfacebetween the slide and the slide cover. The thin film is disposed outsidethe area demarcated by the pattern and preferably spaces the grid fromthe surface of the slide by 10, 20 or 50 μm.

In this way, the slide cover according to the present invention, can beused with standard slides which have no chambers formed thereon.

These and other features of the present invention will be describedhereinafter with reference to the foregoing detailed description takenwith the attached drawings, wherein

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1a is a perspective view of the biological assembly in accordancewith the present invention;

FIG. 1b is a sectional view taken along line Ib--Ib in FIG. 1a;

FIG. 2a is a perspective view of the upper surface of one embodiment ofthe slide cover shown in FIG. 1a;

FIG. 2b is a perspective view of the under surface of one embodiment ofthe slide cover shown in FIG. 1a;

FIG. 3a is a perspective view of an alternative embodiment of the slidecover in accordance with the present invention;

FIG. 3b is a side view of the slide cover of FIG. 3a in use on aconventional slide;

FIG. 3c is a perspective view of another embodiment of the slide cover;and

FIG. 4 is a perspective view of a further embodiment of the presentinvention.

DETAILED DESCRIPTION OF THE INVENTION

As shown in FIGS. 1a and 1b, the biological assembly 1 includes a slide2 and slide covers 3, 3'. The slide 2 includes a substrate 20 preferablymade of glass and which is transparent to allow light to passtherethrough for observation with a microscope. The substrate 20 has atop surface 21 and a bottom surface 22. On top surface 21 is a thin film23 which is opaque and which has a given height, which is preferably 10,20 or 50 μm.

The slide includes at least one sample chamber 24 surrounded by anoverflow chamber 25. The sample chamber 24 is formed by an opening inthe thin film 23 and the overflow chamber 25 is also formed by anopening in the thin film 23The openings are configured to form aseparating wall 26 between the overflow chamber 25 and the samplechamber 24. The separating wall preferably has a width of 200 to 800 μm.In the embodiment shown if FIG. 1a and 1b, the sample chamber ispreferably circular in cross section and thus a cylindrical chamberwhereas the overflow chamber is toroidal in cross section and is coaxialwith the sample chamber. In the embodiment shown in FIG. 1a, two samplechambers and associated overflow chambers are utilized although slidescan be produced with a single sample chamber and overflow chamber orwith more than two, preferably three sample chambers and over flowchambers.

When two or more sample chambers are utilized, a separator 27 isdisposed therebetween and having a height of approximately 100-500 μm.

The thin film 23 is preferably composed of an a photoresist, asdisclosed in U.S. Pat. No. 4,911,782, an epoxy or a hydrophobic ink andis either printed on or patterned by other conventional methods asdisclosed in U.S. Pat. No. 4,911,782.

The biological assembly also includes a slide cover 3, 3', which is usedto cover the sample in the sample chamber 24 during use of the slide ina microscope.

The slide cover 3 comprises a substrate 30, which is preferably made ofa transparent glass 0.1 to 0.5 mm thick and has a top surface 31 and alower surface 32, which is superposed on the upper surface of the thinfilm 23 during use as shown in FIG. 1b. The slide cover also includes apattern of thin film material 36 disposed on the upper surface 31 andhaving a width of 200 to 800 μm. In the embodiment shown in FIG. 1a, thepattern 36 is identical to the separating wall 26 and is used todemarcate an area 37 therewithin and to permit alignment of the area 37with the sample chamber 24 by matching the pattern 36 with the wall 26.

The slide cover 3 also includes a grid of perpendicular lines formed inthe lower surface 32 of the slide cover substrate 30. This grid ispreferably 10 boxes ×10 boxes with each box being 0.1 mm on a side. Thegrid is formed in the surface of the substrate by a laser etching methodcalled excimer laser micromachining, which produces a relatively thingrid lines. The grid formed by this method is almost invisible to thenaked eye.

The slide cover 3' shown in FIG. 1a is identical to the slide cover 3with the exception of the grid, which is not present in the area of 37.When the two slide covers are placed on the slide, the separator 27keeps them apart.

FIGS. 2a and 2b show the slide cover of a FIG. 1a and 1b in more detail.

The slide cover of FIG. 2a and 2b can be used independently of the slide2 shown in FIGS. 1a and 1b and can thus be used with other slides havingpredefined chambers.

The grid is etched into the lower surface of the slide cover to bringthe grid as close as possible to the sample, in order to produce themost accurate results since it substantially eliminates parallax.

In use, the user drops a liquid sample into the sample chamber 24. Anyof the sample which exceeds the volume of the sample chamber 24 willflow into the overflow chamber 25. Thereafter, the slide cover 3 isapplied to the top of the slide by aligning the pattern 36 with theseparating wall 26. When viscous fluids are used, the placing of theslide cover on the slide will force excess fluid into the overflowchamber. The grid 38 is now disposed immediately adjacent to the samplein sample chamber 24 as can be seen in FIG. 1b and the slide isintroduced into a microscope so that the user can count the chamber ofcells within a representative one of the squares of the grid pattern 38.

In an alternative embodiment of the present invention as shown in FIGS.3a and 3b, the slide cover 4 can be used with conventional slides havingno chambers formed thereon. In this embodiment, the substrate 40 has thepattern 46 printed on the upper surface 41 thereof. On the lower surface42, the grid 48 is formed therein in the area 47 demarcated by thepattern 46. Disposed outside of the area 47 are thin film segments 49,which have the function of spacers, and which space the surface 42 ofthe slide cover from the upper surface 6 of a conventional slide 5 shownin FIG. 3b. The spacers 49 are made of thin film and have a thickness ofpreferably 10, 20 or 50 μm. The cover 4 also has indicia 50 indicatingwhich side thereof the grid is on, to aid the user.

As shown in FIG. 3b, a sample S is placed on the surface 6 of a slide 15and the slide cover 4 is disposed thereover with the spacers 49 restingon the surface 6. As a result of the fact that the spacers spaced thesurface 42 a fixed distance from the top surface of the slide, a knownvolume of sample S is disposed within the confines of the grid 48, thusenabling measurements to be made.

In FIG. 3c, the slide cover 4' has the pattern 46' disposed on the lowersurface 42. In this embodiment, the pattern itself can act as a spacerand accordingly, the diameter of the pattern 46' can be made greater toprovide stability, so that the film segment spacers 49 can beeliminated.

In a further embodiment shown in FIG. 4, the biological assemblyincludes a slide 7 and slide covers 8. The slide includes a substrate 72of transparent glass which is preferably 75 mm long and 25 mm wide and 1mm thick. The top surface of substrate 72 is covered with thin film 73having a height of preferably 10, 20 or 50 μm and openings thereindemarcating cylindrical sample chambers 74 and rectangular overflowchambers 75 and forming circular separating walls 76 preferably having awidth of 700 to 800 μm.

The cylindrical sample chambers 74 preferably have a diameter of 16 mm.The rectangular overflow chambers 75 are preferably 23 mm long and 10 mmwide. For a thin film of 20 mm, the sample chamber has a volume of 4 μland the overflow chamber has a volume of 5 μl. The slide covers 8comprise a substrate 81 of transparent glass which has a grid 88 etchedinto the lower surface thereof at a location within the circular pattern86 which is disposed on the upper surface of substrate 81. The substrateis preferably 27 mm long and 22 mm wide and 0.33 mm thick and thepattern 86 and grid 88 are positioned to fall at the center of thesample chamber 74 when the slide cover is disposed on the slide with theedges of the slide cover coaxial with the edges of the sample chamber.

When thin film 73 is hydrophobic, any excess liquid which exceeds theoverflow chamber 75, will be limited in its migration.

It is understood that the present invention is not limited to theparticular embodiments illustrated and described hereinabove but thatvarious changes and modifications may be made without departing from thespirit and scope of the invention as defined in the claims.

What is claimed is:
 1. A biological assembly for effecting theexamination of a sample in a liquid medium under magnification,comprising:a transparent substrate having a first main surface; meansforming at least one sample chamber and an immediately adjacent overflowchamber on the first main surface and receptive of a sample in a liquidmedium, said means comprising a thin film having for each sample chamberand overflow chamber, means defining a first opening thereincorresponding to the sample chamber and means defining a second openingsurrounding the first opening and corresponding to the overflow chamber,wherein the first and second openings form a separating wall of thinfilm therebetween; and a transparent cover having first main surfacesuperposable on the thin film to cover the at least one sample chamberand overflow chamber when a sample is received therein, wherein thecover has a second main surface opposite the first main surface andhaving a thin film pattern thereon demarcating an area for aligning thearea of the cover with the at least one sampling chamber.
 2. Thebiological assembly according to claim 1, wherein the first opening iscircular and the second opening is toroidal.
 3. The biological assemblyaccording to claim 1, wherein the first opening is circular and thesecond opening is rectangular.
 4. The biological assembly according toclaim 1, wherein the thin film has a height of from 10 to 50 μm.
 5. Thebiological assembly according to claim 1, wherein the separating wallhas a width of 200 to 800 μm.
 6. The biological assembly according toclaim 1, wherein said at least one sample chamber has a volume of 2 to15 μl.
 7. The biological assembly according to claim 1, wherein the thinfilm pattern is identical in configuration to said separating wail. 8.The biological assembly according to claim 1, wherein the cover has agrid pattern in the first main surface within the area demarcated bysaid thin film pattern.
 9. The biological assembly according to claim 1,wherein the thin film comprises at least one of a photoresist, an epoxyand a hydrophobic ink.
 10. A biological assembly for effectingexamination of a sample in a liquid medium under magnification,comprising:a transparent substrate having a first main surface; meansforming at least one sample chamber on the first main surface andreceptive of a sample in a liquid medium, said means comprising a thinfilm having for each sample chamber, means defining an opening thereincorresponding to the sample chamber and means defining a open areasurrounding the opening, wherein the opening and open area form aseparating wall of thin film therebetween; and a transparent coverhaving fast main surface superposable on the thin film to cover the atleast one sample chamber when a sample is received therein, wherein thecover has a second main surface opposite the first main surface andhaving a thin film pattern thereon demarcating an area for aligning thearea of the cover with the at least one sampling chamber.
 11. Thebiological assembly according to claim 10, wherein the opening iscircular.
 12. The biological assembly according to claim 10, wherein thethin film has a height of from 10 to 50 μm.
 13. The biological assemblyaccording to claim 10, wherein the separating wall has a width of 200 to800 μm.
 14. The biological assembly according to claim 10, wherein saidat least one sample chamber has a volume of 2 to 15 μl.
 15. Thebiological assembly according to claim 10, wherein the thin film patternis identical in configuration to said separating wail.
 16. Thebiological assembly according to claim 10, wherein the cover has a gridpattern in the first main surface within the area demarcated by saidthin film pattern.
 17. The biological assembly according to claim 10,wherein the thin film comprises at least one of a photoresist, an epoxyand a hydrophobic ink. .Iadd.
 18. A biological assembly for effectingexamination of a sample in a liquid medium under magnification,comprising:a transparent substrate having a surface for receiving thesample; a transparent cover having a first main surface superposed onthe surface of the substrate during use and comprising a grid patternand spacers disposed outwardly of the grid pattern for spacing the firstmain surface apart from the substrate surface by a distance when thecover is superposed on the substrate and a second main surface oppositethe first main surface and having a first thin film pattern thereondemarcating an area including the grid pattern for aligning the gridpattern over the sample when the cover is superposed on the substrate..Iaddend..Iadd.19. The biological assembly according to claim 18,wherein the first thin film pattern is a ring surrounding the gridpattern. .Iaddend..Iadd.20. The biological assembly according to claim18, wherein the spacers comprise a second thin film pattern on the firstmain surface. .Iaddend..Iadd.21. The biological assembly according toclaim 20, wherein the second thin film pattern comprises circles incorners of the cover. .Iaddend..Iadd.22. The biological assemblyaccording to claim 18, wherein the spacers space the first main surfacefrom the substrate surface by 10 microns. .Iaddend..Iadd.23. Thebiological assembly according to claim 18, wherein the spacers space thefirst main surface from the substrate surface by 20 microns..Iaddend..Iadd.24. The biological assembly according to claim 18,wherein the spacers space the first main surface from the substratesurface by 50 microns. .Iaddend..Iadd.25. A biological assembly foreffecting examination of a sample in a liquid medium under magnificationcomprising: a transparent slide having a completely planar surface forreceiving the sample; a transparent slide cover having a first mainsurface superposed on the surface of the slide during use and comprisinga grid pattern, a thin film pattern demarcating an area including thegrid pattern for aligning the grid pattern over the sample when theslide cover is superposed on the slide and a plurality of discretespacers disposed outwardly of and separated from the thin film patternwithout enclosing same for spacing the first main surface apart from theslide surface by a distance when the slide cover is superposed on theslide. .Iaddend..Iadd.26. The biological assembly according to claim 25,wherein the thin film pattern is a ring surrounding the grid pattern..Iaddend..Iadd.27. The biological assembly according to claim 25,wherein the spacers space the first main surface from the slide surfaceby 10 microns. .Iaddend..Iadd.28. The biological assembly according toclaim 25, wherein the spacers space the first main surface from theslide surface by 20 microns. .Iaddend..Iadd.29. The biological assemblyaccording to claim 25, wherein the spacers space the first main surfacefrom the slide surface by 50 microns. .Iaddend.